Stem cells continuously produce highly differentiated but short-lived cells, such as blood, skin, intestinal epithelium, and sperm cells throughout life. Asymmetric stem cell division, which produces one self-renewed stem cell and one differentiated cell, plays a critical role in maintaining tissue homeostasis. The aim of this research proposal is to investigate the molecular and cellular mechanisms that govern stem cell behavior, in particular, the regulation of asymmetric stem cell division, using Drosophila male germ line stem cells (GSCs) as a model system. We have previously shown that centrosome orientation within GSCs with respect to the niche plays a central role in asymmetric GSC division. Our preliminary results suggest the existence of a checkpoint mechanism that monitors correct centrosome orientation within GSCs, ensuring an asymmetric outcome of stem cell division. We will characterize this checkpoint (referred to as the orientation checkpoint) and aim to identify the components of the checkpoint mechanism. We will investigate at which cell cycle stage the checkpoint arrests/delays the GSC cell cycle in response to centrosome misorientation. We will also take a candidate approach to identify the components of the orientation checkpoint. Furthermore, we will investigate the potential role of this checkpoint mechanism in the age-related decline in tissue regenerative capacity. We aim to understand the mechanisms by which adult stem cells ensure to maintain both stem cell and differentiated cell populations. Understanding of such mechanisms may allow us to manipulate stem cell behavior; for example, normal stem cells can be expanded in culture for transplantation, or we can inhibit the expansion of cancerous stem cells.